Immunologic memory is the defining characteristic of the adaptive immune response. Studies in experimental animals and observations in humans indicate that memory T lymphocytes are critical for protection from many pathogens, particularly viruses, and that these cells are maintained for long periods of time in vivo. We hypothesize that the human virus-specific memory T lymphocyte repertoire undergoes quantitative, and also qualitative, changes over time, consistent with the model of cross-reactive activation of memory T cells by unrelated antigens. We will address this hypothesis through the detailed analysis of the human vaccinia virus-specific T cell repertoire over time. The Specific Aims are: 1) To determine whether the vaccinia virus-specific memory T cell repertoire changes during the induction and maintenance of memory T cells over the first 4 years after vaccination, by analyzing the frequency and T cell receptor (TCR) spectratype of virus- and peptide-specific T cells in serial PBMC obtained up to 4 years after vaccination. 2) To determine whether there are significant differences in the long-term vaccinia virus-specific memory T cell repertoire in healthy adults related to age and time since vaccination, by analyzing the frequency and TCR spectratype of virus- and peptide-specific T cells in PBMC and their associations with the age of subjects and time after vaccination. 3) To determine whether human virus-specific memory T cells are maintained because of periodic activation by exposure to unrelated antigens, by analyzing- a) the expression of activation markers on memory vaccinia virus-specific T cells, b) whether residual vaccinia viral DNA is present in autopsy lymph node tissues from persons vaccinated during childhood, and c) whether immunization with vaccinia virus induces changes in the frequency and TCR-spectratype of memory influenza- or EBV-specific T cells. Information obtained from this study will improve the understanding of vaccine efficacy and aged related susceptibility to infection. This project directly supports the overall objective of this Program Project by addressing the induction and maintenance of virus-specific memory T lymphocytes.